scholarly journals Sleeping ribosomes: bacterial signaling triggers RaiA mediated persistence to aminoglycosides

2020 ◽  
Author(s):  
Manon Lang ◽  
Evelyne Krin ◽  
Chloé Korlowski ◽  
Odile Sismeiro ◽  
Hugo Varet ◽  
...  
Keyword(s):  
Sucrose Gradient ◽  
Gradient Analysis ◽  
Cell Formation ◽  
Fold Increase ◽  
Growth Conditions ◽  
Lag Phase ◽  
Bacterial Signaling ◽  
Tryptophan Degradation ◽  
70S Ribosome ◽  
Persistent Cell

AbstractIndole is a small molecule derived from tryptophan degradation and proposed to be involved in bacterial signaling. We find that indole secretion is induced by sublethal tobramycin concentrations and increases persistence to aminoglycosides in V. cholerae. Indole transcriptomics showed strongly increased expression of raiA, a ribosome associated factor. Deletion of raiA abolishes the appearance of indole dependent persisters to aminoglycosides, while its overexpression leads to 100-fold increase of persisters, and a reduction in lag phase, evocative of increased active 70S ribosome content, which was confirmed by sucrose gradient analysis. We propose that, under stress conditions, inactive 70S ribosomes are associated with RaiA to be stored and rapidly reactivated when growth conditions become favorable again, in a mechanism different than ribosome hibernation. Our results point to an active process of persistent cell formation, through ribosome protection during translational stress and relief upon antibiotic removal. Translation is a universal process, and these results could help elucidate a mechanism of persistence formation in a controlled, thus inducible way.

scholarly journals Low Glucose Mediated Fluconazole Tolerance in Cryptococcus neoformans

2021 ◽  
Vol 7 (6) ◽  
pp. 489
Author(s):  
Somanon Bhattacharya ◽  
Natalia Kronbauer Oliveira ◽  
Anne G. Savitt ◽  
Vanessa K. A. Silva ◽  
Rachel B. Krausert ◽  
...  
Keyword(s):  
Cryptococcus Neoformans ◽  
Mitochondrial Function ◽  
Efflux Pump ◽  
Nile Red ◽  
Fold Increase ◽  
Growth Conditions ◽  
Chromosome 1 ◽  
Normal Glucose ◽  
Low Glucose ◽  
Synthetic Media

Chronic meningoencephalitis is caused by Cryptococcus neoformans and is treated in many parts of the world with fluconazole (FLC) monotherapy, which is associated with treatment failure and poor outcome. In the host, C. neoformans propagates predominantly under low glucose growth conditions. We investigated whether low glucose, mimicked by growing in synthetic media (SM) with 0.05% glucose (SMlowglu), affects FLC-resistance. A > 4-fold increase in FLC tolerance was observed in seven C. neoformans strains when minimum inhibitory concentration (MIC) was determined in SMlowglu compared to MIC in SM with normal (2%) glucose (SMnlglu). In SMlowglu, C. neoformans cells exhibited upregulation of efflux pump genes AFR1 (8.7-fold) and AFR2 (2.5-fold), as well as decreased accumulation (2.6-fold) of Nile Red, an efflux pump substrate. Elevated intracellular ATP levels (3.2-fold and 3.4-fold), as well as decreased mitochondrial reactive oxygen species levels (12.8-fold and 17-fold), were found in the presence and absence of FLC, indicating that low glucose altered mitochondrial function. Fluorescence microscopy revealed that mitochondria of C. neoformans grown in SMlowglu were fragmented, whereas normal glucose promoted a reticular network of mitochondria. Although mitochondrial membrane potential (MMP) was not markedly affected in SMlowglu, it significantly decreased in the presence of FLC (12.5-fold) in SMnlglu, but remained stable in SMlowglu-growing C. neoformans cells. Our data demonstrate that increased FLC tolerance in low glucose-growing C. neoformans is the result of increased efflux pump activities and altered mitochondrial function, which is more preserved in SMlowglu. This mechanism of resistance is different from FLC heteroresistance, which is associated with aneuploidy of chromosome 1 (Chr1).

Effect of previous cutting interval and of leaf area remaining after cutting on regrowth of Macroptilium atropurpureum cv. Siratro

1974 ◽  
Vol 14 (68) ◽  
pp. 343 ◽  
Author(s):  
RJ Jones
Keyword(s):  
Leaf Area ◽  
Practical Implication ◽  
Soluble Sugars ◽  
Ground Level ◽  
Fold Increase ◽  
Water Soluble ◽  
Hot Water ◽  
Lag Phase ◽  
Area Index ◽  
Plant Yield

Experiments with Siratro were conducted at Samford, south east Queensland to study the effects of previous cutting and defoliation treatments on regrowth. In the first experiment, swards of Siratro were cut at 7.5 cm above ground level every 4 weeks, every 8 weeks or cut once at 16 weeks during spring and summer. Regrowth of all treatments over ten weeks was measured after varying (by leaf removal) the stubble leaf area index (LAI) of the plots cut every four weeks. Pattern of regrowth yield was similar for all treatments with a pronounced lag phase after cutting. Regrowth yield after 10 weeks differed between treatments and was linearly related (P < 0.01 ) to residual LAI in the stubble at the start of regrowth. In the absence of stubble leaves, plots previously cut at 16 weeks or at 8 weeks yielded marginally more than those cut every 4 weeks. There were no marked treatment differences in gross root morphology other than a two fold increase in stolon rooting for the 16-week treatment. Nitrogen content of the roots (mean 1.38 per cent) was unaffected by treatment, but the per cent hot water soluble sugars were lower for the 16 week defoliation treatment than for the 8-week and the 4-week treatments. In the second experiment individual plants were cut to a uniform stubble every 4 weeks and either 0, 5, or 10 leaves were left. Dry weight of regrowth and stolon development were greatest when most leaves were left. Two thirds of the plants died after six cuttings with complete defoliation but none died when either 5 or 10 leaves were retained. Plant survival was not related to plant yield or degree of stoloniferous development. However, there was a strong correlation between stolon number and plant yield under this intensive cutting regime. The practical implication of the results in the management of Siratro is discussed.

Regulation of the utilization of mRNA for eucaryotic elongation factor Tu in Friend erythroleukemia cells

1987 ◽  
Vol 7 (2) ◽  
pp. 687-697
Author(s):  
T R Rao ◽  
L I Slobin
Keyword(s):  
Stationary Phase ◽  
Elongation Factor ◽  
Fold Increase ◽  
Growth Conditions ◽  
Fresh Medium ◽  
Initiation Factor ◽  
Elongation Factor Tu ◽  
Erythroleukemia Cells ◽  
Friend Erythroleukemia Cells

When Friend erythroleukemia cells were allowed to grow to stationary phase (2 X 10(6) to 3 X 10(6) cells per ml), approximately 60% of the mRNA for eucaryotic elongation factor Tu (eEF-Tu) sedimented at less than or equal to 80S, and most of the remaining factor mRNA was associated with small polysomes. Under the same growth conditions, greater than 90% of the mRNA for eucaryotic initiation factor 4A remained associated with polysomes. The association of eEF-Tu mRNA with polysomes changed dramatically when stationary-phase cells were treated with fresh medium. After 1 h in fresh medium, approximately 90% of eEF-Tu mRNA in Friend cells was found in heavy polysomes. Associated with the shift of eEF-Tu mRNA into heavy polysomes, we found at least a 2.6-fold increase in the synthesis of eEF-Tu in vivo as well as a remarkable 40% decrease in the total amount of eEF-Tu mRNA per cell. Our data raise the possibility that eEF-Tu mRNA that has accumulated in ribonucleoprotein particles in stationary-phase cells is degraded rather than reutilized for eEF-Tu synthesis.

scholarly journals Identification and Characterization ofMAE1, the Saccharomyces cerevisiae Structural Gene Encoding Mitochondrial Malic Enzyme

1998 ◽  
Vol 180 (11) ◽  
pp. 2875-2882 ◽  
Author(s):  
Eckhard Boles ◽  
Patricia de Jong-Gubbels ◽  
Jack T. Pronk
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Enzyme Activity ◽  
Pyruvate Kinase ◽  
Malic Enzyme ◽  
Fold Increase ◽  
Growth Conditions ◽  
Anaerobic Growth ◽  
Oxidative Decarboxylation ◽  
Mrna Levels ◽  
Malic Enzyme Activity

ABSTRACT Pyruvate, a precursor for several amino acids, can be synthesized from phosphoenolpyruvate by pyruvate kinase. Nevertheless, pyk1 pyk2 mutants of Saccharomyces cerevisiae devoid of pyruvate kinase activity grew normally on ethanol in defined media, indicating the presence of an alternative route for pyruvate synthesis. A candidate for this role is malic enzyme, which catalyzes the oxidative decarboxylation of malate to pyruvate. Disruption of open reading frame YKL029c, which is homologous to malic enzyme genes from other organisms, abolished malic enzyme activity in extracts of glucose-grown cells. Conversely, overexpression ofYKL029c/MAE1 from the MET25 promoter resulted in an up to 33-fold increase of malic enzyme activity. Growth studies with mutants demonstrated that presence of either Pyk1p or Mae1p is required for growth on ethanol. Mutants lacking both enzymes could be rescued by addition of alanine or pyruvate to ethanol cultures. Disruption of MAE1 alone did not result in a clear phenotype. Regulation of MAE1 was studied by determining enzyme activities and MAE1 mRNA levels in wild-type cultures and by measuring β-galactosidase activities in a strain carrying a MAE1::lacZ fusion. Both in shake flask cultures and in carbon-limited chemostat cultures,MAE1 was constitutively expressed. A three- to fourfold induction was observed during anaerobic growth on glucose. Subcellular fractionation experiments indicated that malic enzyme in S. cerevisiae is a mitochondrial enzyme. Its regulation and localization suggest a role in the provision of intramitochondrial NADPH or pyruvate under anaerobic growth conditions. However, since null mutants could still grow anaerobically, this function is apparently not essential.

Regulation of the Synthesis of Ribulose-l,5-bisphosphate Carboxylase and Its Subunits in the Flagellate Chlorogonium elongatum. II. Coordinated Synthesis of the Large and Small Subunits

1981 ◽  
Vol 36 (11-12) ◽  
pp. 942-950 ◽  
Author(s):  
Peter Westhoff ◽  
Kurt Zimmermann ◽  
Frank Boege ◽  
Klaus Zetsche
Keyword(s):  
Rna Synthesis ◽  
De Novo ◽  
Fold Increase ◽  
Growth Conditions ◽  
Autotrophic Growth ◽  
De Novo Synthesis ◽  
Bisphosphate Carboxylase ◽  
Unicellular Green Alga ◽  
Protein And Rna Synthesis ◽  
Ribulosebisphosphate Carboxylase

Abstract Transfer of heterotrophically grown cells of the unicellular green alga Chlorogonium elongatum to autotrophic growth conditions causes a 10 -15 fold increase in the amount of the chloroplastic enzyme ribulose-1,5-bisphosphate carboxylase. This increase was found to be due to de novo synthesis. The relative proportions of large and small subunits of the enzyme do not change. Their ratio is close to 3.4, the proportions in weight of the two subunits in the holoenzyme. Continous labelling with [35S]sulfate reveals that the ratios of incorporation into large and small subunits are essentially the same in autotrophic and heterotrophic cells. Pulse-chase experiments show that the subunits are degraded synchronously. The coordinated subunit synthesis cannot be uncoupled using inhibitors of protein and RNA synthesis or high temperature of cultivation of the alga. The results suggests a very tightly coordinated synthesis of the large and small subunits of ribulosebisphosphate carboxylase.

GROWTH STUDIES ON ARTHROBACTER GLOBIFORMIS: II. CHANGES IN MACROMOLECULAR LEVELS DURING GROWTH

1962 ◽  
Vol 8 (5) ◽  
pp. 655-661 ◽  
Author(s):  
I. L. Stevenson
Keyword(s):  
Stationary Phase ◽  
Fold Increase ◽  
Growth Cycle ◽  
Phase Cell ◽  
Lag Phase ◽  
Differential Rate ◽  
Protein Ratio ◽  
Stationary Phase Culture ◽  
Protein Levels ◽  
Dna And Protein Synthesis

Changes in macromolecular levels (RNA, DNA, protein) have been followed during the growth cycle of A. globiformis. When a stationary phase culture is transferred to fresh medium a 12-fold increase in RNA level and 6-fold increases in DNA and protein levels are observed during the predivisional lag phase. Initially RNA synthesis precedes DNA and protein synthesis but all reach the same differential rate 2 to 3 hours prior to division. During the predivisional lag period the RNA/protein ratio per cell expands from 0.19 to 0.36. Once division occurs, cells of A. globiformis remain in the enlarged pleomorphic form until the medium becomes limiting; at this time synthesis of macromolecules ceases and the continued division (three to four generations) results in progressively smaller cells until the coccoid stationary phase cell-type is reached.

Hemoglobin Biosynthesis in Vitreoscilla stercoraria DW: Cloning, Expression, and Characterization of a New Homolog of a Bacterial Globin Gene

1998 ◽  
Vol 64 (6) ◽  
pp. 2220-2228 ◽  
Author(s):  
Meenal Joshi ◽  
Shekhar Mande ◽  
Kanak L. Dikshit
Keyword(s):  
Globin Gene ◽  
Three Dimensional ◽  
Fold Increase ◽  
Binding Pocket ◽  
Growth Conditions ◽  
Dimensional Structure ◽  
Strictly Aerobic ◽  
Coding Region ◽  
Oxygen Control ◽  
Constitutive Production

ABSTRACT In the strictly aerobic, gram-negative bacteriumVitreoscilla strain C1, oxygen-limited growth conditions create a more than 50-fold increase in the expression of a homodimeric heme protein which was recognized as the first bacterial hemoglobin (Hb). The recently determined crystal structure ofVitreoscilla Hb has indicated that the heme pocket of microbial globins differs from that of eukaryotic Hbs. In an attempt to understand the diverse functions of Hb-like proteins in prokaryotes, we have cloned and characterized the gene (vgb) encoding an Hb-like protein from another strain of Vitreoscilla,V. stercoraria DW. Several silent changes were observed within the coding region of the V. stercoraria vgb gene. Apart from that, V. stercoraria Hb exhibited interesting differences between the A and E helices. Compared to its Hb counterpart from Vitreoscilla strain C1, the purified preparation ofV. stercoraria Hb displays a slower autooxidation rate. The differences between Vitreoscilla Hb and V. stercoraria Hb were mapped onto the three-dimensional structure of Vitreoscilla Hb, which indicated that the four changes, namely, Ile7Val, Ile9Thr, Ile10Ser, and Leu62Val, present within theV. stercoraria Hb fall in the region where the A and E helices contact each other. Therefore, alteration in the relative orientation of the A and E helices and the corresponding conformational change in the heme binding pocket of V. stercoraria Hb can be correlated to its slower autooxidation rate. In sharp contrast to the oxygen-regulated biosynthesis of Hb in Vitreoscillastrain C1, production of Hb in V. stercoraria has been found to be low and independent of oxygen control, which is supported by the absence of a fumarate and nitrate reductase regulator box within the V. stercoraria vgb promoter region. Thus, the regulation mechanisms of the Hb-encoding gene appear to be quite different in the two closely related species ofVitreoscilla. The relatively slower autooxidation rate ofV. stercoraria Hb, lack of oxygen sensitivity, and constitutive production of Hb suggest that it may have some other function(s) in the cellular physiology of V. stercorariaDW, together with facilitated oxygen transport, predicted for earlier reported Vitreoscilla Hb.

Mechanisms of Bacterial Survival in Chlorinated Drinking Water

1988 ◽  
Vol 20 (11-12) ◽  
pp. 145-151 ◽  
Author(s):  
Mark W. LeChevallier ◽  
Cheryl D. Cawthon ◽  
Ramon G. Lee
Keyword(s):  
Drinking Water ◽  
Resistance Mechanism ◽  
Fold Increase ◽  
Growth Conditions ◽  
Resistance Mechanisms ◽  
Bacterial Survival ◽  
Nutrient Effects ◽  
Chlorinated Drinking Water ◽  
Bacterial Encapsulation ◽  
Microscope Slides

Experiments showed that attachment of bacteria to surfaces provided the greatest increase in disinfection resistance. Attachment of high nutrient grown, unencapsulated, Klebsiellapneumoniae to glass microscope slides afforded the microorganisms as much as a 150 fold increase in disinfection resistance. Other mechanisms which increased disinfection resistance included: the age of the biofilm, bacterial encapsulation and previous growth conditions (e.g. growth medium, and growth temperature). These factors increased chlorine resistance from two to ten fold. The choice of disinfectant residual was shown to influence the type of resistance mechanism observed. Disinfection by free chlorine was affected by surfaces, age of the biofilm, encapsulation and nutrient effects. Disinfection by monochloramine, however, was only affected by surfaces. Importantly, the research showed that these resistance mechanisms were multiplicative (e.g. the resistance provided by one mechanism could be multiplied by the resistance provided by a second). These results provide important insights to understand the survival of bacteria in chlorinated drinking water supplies.

Validation of Predictive Mathematical Models Describing Growth of Staphylococcus aureus

1996 ◽  
Vol 59 (1) ◽  
pp. 11-15 ◽  
Author(s):  
ISABEL WALLS ◽  
VIRGINIA N. SCOTT ◽  
DANE T. BERNARD
Keyword(s):  
Staphylococcus Aureus ◽  
Mathematical Models ◽  
Growth Kinetics ◽  
Growth Rates ◽  
Close Agreement ◽  
Growth Conditions ◽  
Lag Phase ◽  
Growth Data ◽  
Gompertz Equation ◽  
Ph Range

An investigation was performed on the growth of Staphylococcus aureus in a commercially available, sterile, homogeneous food at 12°C with 1.2 and 5.9% NaCl; at 25°C with 10.4% NaCl; and at 20 and 35°C with 1.2, 5.3, 12.5, and 15.8% NaCl; over a pH range of 5.5 to 7.5. Growth data were fitted to the Gompertz equation and the resulting growth kinetics were compared with predictions from the Pathogen Modeling Program (PMP) and Food MicroModel (FMM). For the PMP, predicted lag-phase durations varied from 0.5 to 130 h longer than the observed values. In general, close agreement with growth rates was obtained but there was a 10-fold difference in one case. For FMM, predicted lag-phase durations ranged from 27 h shorter to 47 h longer than the observed values. Again, close agreement with growth rates was obtained, but in one case a fivefold difference was observed. In general, for the sterile foods used under the growth conditions tested, the models underestimated the growth of S. aureus. This implies that while the models can be used as a guide to indicate growth rates in foods they should not be relied upon as the sole determinant of the product's safety.

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